scholarly journals MDM2 antagonists induce p53-dependent apoptosis in AML: implications for leukemia therapy

Blood ◽  
2005 ◽  
Vol 106 (9) ◽  
pp. 3150-3159 ◽  
Author(s):  
Kensuke Kojima ◽  
Marina Konopleva ◽  
Ismael J. Samudio ◽  
Masato Shikami ◽  
Maria Cabreira-Hansen ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Transcriptional Activation ◽  
P53 Protein ◽  
Negative Regulator ◽  
Wild Type ◽  
Protein Levels ◽  
P53 Signaling ◽  
P53 Activation ◽  
Induction Of Apoptosis ◽  
Wild Type P53

AbstractAlthough TP53 mutations are rare in acute myeloid leukemia (AML), inactivation of wild-type p53 protein frequently occurs through overexpression of its negative regulator MDM2 (murine double minute 2). Recently, small-molecule antagonists of MDM2, Nutlins, have been developed that inhibit the p53-MDM2 interaction and activate p53 signaling. Here, we study the effects of p53 activation by Nutlin-3 in AML cells. Treatment with MDM2 inhibitor triggered several molecular events consistent with induction of apoptosis: loss of mitochondrial membrane potential, caspase activation, phosphatidylserine externalization, and DNA fragmentation. There was a positive correlation in primary AML samples with wild-type p53 between baseline MDM2 protein levels and apoptosis induced by MDM2 inhibition. No induction of apoptosis was observed in AML samples harboring mutant p53. Colony formation of AML progenitors was inhibited in a dose-dependent fashion, whereas normal CD34+ progenitor cells were less affected. Mechanistic studies suggested that Nutlin-induced apoptosis was mediated by both transcriptional activation of proapoptotic Bcl-2 family proteins, and transcription-independent mitochondrial permeabilization resulting from mitochondrial p53 translocation. MDM2 inhibition synergistically enhanced cytotoxicity of cytosine arabinoside and doxorubicin in AML blasts but not in normal hematopoietic progenitor cells. p53 activation by targeting the p53-MDM2 interaction might offer a novel therapeutic strategy for AML that retain wild-type p53.

scholarly journals DNA strand breaks: the DNA template alterations that trigger p53-dependent DNA damage response pathways.

1994 ◽  
Vol 14 (3) ◽  
pp. 1815-1823 ◽  
Author(s):  
W G Nelson ◽  
M B Kastan
Keyword(s):  
Excision Repair ◽  
P53 Protein ◽  
Dna Strand Breaks ◽  
Wild Type ◽  
Strand Breaks ◽  
Protein Levels ◽  
Dna Damaging Agents ◽  
Wild Type P53 ◽  
Dna Strand ◽  
Dna Template

The tumor suppressor protein p53 serves as a critical regulator of a G1 cell cycle checkpoint and of apoptosis following exposure of cells to DNA-damaging agents. The mechanism by which DNA-damaging agents elevate p53 protein levels to trigger G1/S arrest or cell death remains to be elucidated. In fact, whether damage to the DNA template itself participates in transducing the signal leading to p53 induction has not yet been demonstrated. We exposed human cell lines containing wild-type p53 alleles to several different DNA-damaging agents and found that agents which rapidly induce DNA strand breaks, such as ionizing radiation, bleomycin, and DNA topoisomerase-targeted drugs, rapidly triggered p53 protein elevations. In addition, we determined that camptothecin-stimulated trapping of topoisomerase I-DNA complexes was not sufficient to elevate p53 protein levels; rather, replication-associated DNA strand breaks were required. Furthermore, treatment of cells with the antimetabolite N(phosphonoacetyl)-L-aspartate (PALA) did not cause rapid p53 protein increases but resulted in delayed increases in p53 protein levels temporally correlated with the appearance of DNA strand breaks. Finally, we concluded that DNA strand breaks were sufficient for initiating p53-dependent signal transduction after finding that introduction of nucleases into cells by electroporation stimulated rapid p53 protein elevations. While DNA strand breaks appeared to be capable of triggering p53 induction, DNA lesions other than strand breaks did not. Exposure of normal cells and excision repair-deficient xeroderma pigmentosum cells to low doses of UV light, under conditions in which thymine dimers appear but DNA replication-associated strand breaks were prevented, resulted in p53 induction attributable to DNA strand breaks associated with excision repair. Our data indicate that DNA strand breaks are sufficient and probably necessary for p53 induction in cells with wild-type p53 alleles exposed to DNA-damaging agents.

MJ05, a Novel p53 Activating Compound, Effectively and Selectively Eliminates Human CML Stem/Progenitor Cells

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 1464-1464
Author(s):  
Su Chu ◽  
Catherine Drummond ◽  
Ling Li ◽  
YinWei Ho ◽  
Sonia Lain ◽  
...  
Keyword(s):  
Stem Cells ◽  
Progenitor Cells ◽  
Kinase Inhibitors ◽  
Conflicts Of Interest ◽  
P53 Protein ◽  
Wild Type ◽  
Inhibition Of Proliferation ◽  
Protein Levels ◽  
Cd34 Cells

Abstract Tyrosine kinase inhibitors (TKI) are the mainstay of CML treatment but fail to eliminate leukemia stem cells (LSC), leading to high risk of disease recurrence when treatment is stopped. There is considerable interest in developing new strategies to target CML LSC. We have previously shown that p53 activation following SIRT1 deacetylase inhibition can inhibit growth and survival of TKI-treated CML LSC (Cancer Cell 2012, 21:266). We are therefore interested in investigating other strategies to activate p53 as potential approaches to target CML LSC. While conducting a screen of 20,000 small molecules for ability to activate p53-dependent transcription in TP53 wild-type ARN8 human melanoma cells we identified MJ05 amongst the top-ranking compounds. Importantly MJ05 did not activate p53-dependent transcription in T22 fibroblast cells. Increased p53 protein levels in ARN8 cells were seen within 6 hours and were accompanied by an increase in p21, pig3 and mdm2 mRNA and protein levels. Dependency on p53 was confirmed using p53-null and wild-type H1299 cells. Activation of p53 occurred without concurrent increase in DNA damage evidence by g-H2AX labeling, increased p53 Ser 15-phosphorylation, or inhibition of p53-HDM2 interaction. MJ05 treatment inhibited S-Phase progression of ARN8 cells without inhibition of ATM, ATR or DNA-PK phosphorylation, suggesting a unique mechanism of action. We tested the effect of MJ05 on primary normal or CML CD34+ cell by itself and in combination with TKI inhibitor Nilotinib, and compared with effects on Nutlin, a well studied inhibitor of p53-HDM2 interactions. Apoptosis was assessed by Annexin V labeling, proliferation by CFSE labeling, and colony forming cell (CFC) frequency, in methylcellulose progenitor assays. Treatment with MJ05 (5 and 10μM), with or without Nilotinib (1μM), for 72 hours in the presence of low concentrations of growth factor significantly and selectively increased apoptosis, inhibited proliferation and reduced colony CFC frequency in CML CD34+ cells compared to normal CD34+ cells. Combination of MJ05 with Nilotinib (1μM) resulted in significant increase in apoptosis of CML but not normal CD34+ cells. In contrast treatment with Nutlin (2 and 5 μM) resulted in similar increase in apoptosis in CML and normal CD34+ cells. We next evaluated the effect of treatment with MJ05 (10μM), Nilotinib and the combination for 72 hours on purified CML and normal CD34+CD38- stem/primitive progenitor cells and CD34+CD38+ committed progenitor cells. MJ05 significantly enhanced apoptosis of CML but not normal CD34+CD38- cells and CD34+CD38+ cells. Apoptosis was further enhanced by combination with Nilotinib (Table). MJ05 also resulted in significant reduction of proliferation in CML CD34+38+ and CD34+38- cells, with significantly less inhibition of proliferation of normal cells. MJ05 treatment markedly reduced CFU-GM and BFU-E generation from CML compared to normal CD34+CD38- and CD34+CD38+ cells. The combination with Nilotinib resulted in almost complete abrogation of CML CFC (Table). MJ05 resulted in significantly less inhibition of normal CFC, with greater effect on normal BFU-E compared to CFU-GM. Ongoing xenograft experiments are testing the effect of in vitro treatment with MJ05, Nilotinib or the combination on engraftment of CML and normal stem cells in NSG mice. Our studies indicate that MJ05, a unique, potent and selective p53 activating compound, is remarkably effective in inducing apoptosis and inhibiting growth of primitive CML stem/progenitor cells by itself and to an even greater extent in combination with Nilotinib. MJ05 treatment has significantly lesser effects on normal stem cells, and may offer a promising approach to selectively target CML LSC in combination with Nilotinib.Table 1NormalCMLUntreatedMj05+NilMj05+NilUntreatedMj05NilMj05+NilCD34+38+% Apoptosis4.5±0.19.0±1.14.13±018.8±7.17.5±0.815.9±4.412.3±1.723±7.4CFU-GM86±757.3±4.191±3.852±278.7±19.95±336±141±0BFU-E166±15.315±3.2155.7±9.314±1.595±19.30.7±0.721.3±5.50.3±03CD34+38-% Apoptosis4.2±0.26.1±0.34.2±0.15.3±0.423.0±1.230±4.114.6±3.850.1±7.2CFU-GM67.3±15.941.7±5.549±4.730.3±4.9192.7±50.66±2.372.67±48.70.7±0.7BFU-E58.7±20.211±2.526.3±2.97.6±0.9150.3±58.71±0.672±440.3±0.3 Disclosures: No relevant conflicts of interest to declare.

scholarly journals Interferon-Inducible Protein IFIXα1 Functions as a Negative Regulator of HDM2

2006 ◽  
Vol 26 (5) ◽  
pp. 1979-1996 ◽  
Author(s):  
Yi Ding ◽  
Jin-Fong Lee ◽  
Hua Lu ◽  
Mong-Hong Lee ◽  
Duen-Hwa Yan
Keyword(s):  
Amino Acid ◽  
Gene Family ◽  
Tumor Suppression ◽  
P53 Protein ◽  
Negative Regulator ◽  
Breast Cancer Cell Lines ◽  
Homologous Proteins ◽  
Protein Levels ◽  
P53 Activation ◽  
Inducible Protein

ABSTRACT The 200-amino-acid repeat (HIN-200) gene family with the hematopoietic interferon (IFN)-inducible nuclear protein encodes highly homologous proteins involved in cell growth, differentiation, autoimmunity, and tumor suppression. IFIX is the newest member of the human HIN-200 family and is often downregulated in breast tumors and breast cancer cell lines. The expression of the longest isoform of IFIX gene products, IFIXα1, is associated with growth inhibition, suppression of transformation, and tumorigenesis. However, the mechanism underlying the tumor suppression activity of IFIXα1 is not well understood. Here, we show that IFIXα1 downregulates HDM2, a principal negative regulator of p53, at the posttranslational level. IFIXα1 destabilizes HDM2 protein and promotes its ubiquitination. The E3 ligase activity of HDM2 appears to be required for this IFIXα1 effect. Importantly, HDM2 downregulation is required for the IFIXα1-mediated increase of p53 protein levels, transcriptional activity, and nuclear localization, suggesting that IFIXα1 positively regulates p53 by acting as a negative regulator of HDM2. We found that IFIXα1 interacts with HDM2. Interestingly, the signature motif of the HIN-200 gene family, i.e., the 200-amino-acid HIN domain of IFIXα1, is sufficient not only for binding HDM2 but also for downregulating it, leading to p53 activation. Finally, we show that IFIX mediates HDM2 downregulation in an IFN-inducible system. Together, these results suggest that IFIXα1 functions as a tumor suppressor by repressing HDM2 function.

scholarly journals The correlation between the use of personal protective equipment and level wild-type p53 of dental technicians in Surabaya

2017 ◽  
Vol 50 (1) ◽  
pp. 19 ◽  
Author(s):  
Puspa Dila Rohmaniar ◽  
Titiek Berniyanti ◽  
Retno Pudji Rahayu
Keyword(s):  
Personal Protective Equipment ◽  
P53 Protein ◽  
Pearson Correlation ◽  
P53 Gene ◽  
Working Environment ◽  
Protective Equipment ◽  
Wild Type ◽  
Cross Sectional ◽  
Protein Levels ◽  
Wild Type P53

Background: Exposure of metals among dental technicians that come from the working environment can lead to the formation reactive oxygen species (ROS). ROS can cause mutations in the p53 gene (p53). The mutation is transversion mutation GuanineThymine. p53 mutations can lead to low expression of the wild-type p53 protein (p53). Wild-type p53 involved in many biological processes such as regulation of genes involved in cell cycle, cell growth after DNA damage, and apoptosis. However, exposure to metals among dental technicians can be prevented through the use of personal protective equipment (PPE) during work. Purpose: The purpose of this study was to analyze the correlation between the use of personal protective equipment to wild-type p53 protein levels among dental technicians in Surabaya. Method: This study was observational analytic with cross sectional approach. 40 samples were taken by random sampling. Data were retrieved through interviews and observations. Wild-type p53 was analyzed from saliva with indirect ELISA method. Analysis of data used Kolmogorov Smirnov normality test and a Pearson correlation test. Value significance was p<0.05 (95% confidence level). Result: There was a significant association between the use of personal protective equipment with wild-type p53 levels with p=0.002 Conclusion: The use PPE properly is positively correlated with the wild-type p53 protein levels of dental technicians in Surabaya.

Physical and functional interaction between wild-type p53 and mdm2 proteins

1994 ◽  
Vol 14 (2) ◽  
pp. 1171-1178
Author(s):  
D S Haines ◽  
J E Landers ◽  
L J Engle ◽  
D L George
Keyword(s):  
Transcriptional Activation ◽  
Control Function ◽  
P53 Protein ◽  
Growth Control ◽  
Activation Function ◽  
Protein Domain ◽  
Wild Type ◽  
Mdm2 Gene ◽  
Transient Transfection Assay ◽  
Wild Type P53

The mdm2 oncogene, which is often amplified in mammalian tumors, produces a number of transcripts that encode distinct protein forms. Previous studies demonstrating that overexpression of the mdm2 gene can activate its transforming potential, and can inhibit the transcriptional activation function of p53, prompted us to begin to explore possible functional differences among the various mdm2 products. Utilizing a transient transfection assay, we have evaluated four naturally occurring murine mdm2 forms for their ability to inhibit p53-mediated transcriptional activation of reporter genes regulated by p53 response elements. Three of these mdm2 forms were found to physically associate with the wild-type p53 protein and to possess the ability to inhibit its transactivation function. A fourth form failed to exhibit either of these functions. This last mdm2 form lacks the N-terminal protein domain that is present in the other three splice forms examined, pointing to this region as one that is critical for complex formation with the p53 protein. Identifying such differences among mdm2 proteins provides important clues for dissecting their functional domains, and emphasizes that defining the individual properties of these products will be critical in elucidating the overall growth control function of the mdm2 gene.

scholarly journals Physical and functional interaction between wild-type p53 and mdm2 proteins.

1994 ◽  
Vol 14 (2) ◽  
pp. 1171-1178 ◽  
Author(s):  
D S Haines ◽  
J E Landers ◽  
L J Engle ◽  
D L George
Keyword(s):  
Transcriptional Activation ◽  
Control Function ◽  
P53 Protein ◽  
Growth Control ◽  
Activation Function ◽  
Protein Domain ◽  
Wild Type ◽  
Mdm2 Gene ◽  
Transient Transfection Assay ◽  
Wild Type P53

The mdm2 oncogene, which is often amplified in mammalian tumors, produces a number of transcripts that encode distinct protein forms. Previous studies demonstrating that overexpression of the mdm2 gene can activate its transforming potential, and can inhibit the transcriptional activation function of p53, prompted us to begin to explore possible functional differences among the various mdm2 products. Utilizing a transient transfection assay, we have evaluated four naturally occurring murine mdm2 forms for their ability to inhibit p53-mediated transcriptional activation of reporter genes regulated by p53 response elements. Three of these mdm2 forms were found to physically associate with the wild-type p53 protein and to possess the ability to inhibit its transactivation function. A fourth form failed to exhibit either of these functions. This last mdm2 form lacks the N-terminal protein domain that is present in the other three splice forms examined, pointing to this region as one that is critical for complex formation with the p53 protein. Identifying such differences among mdm2 proteins provides important clues for dissecting their functional domains, and emphasizes that defining the individual properties of these products will be critical in elucidating the overall growth control function of the mdm2 gene.

scholarly journals p53 dynamics vary between tissues and are linked with radiation sensitivity

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Jacob Stewart-Ornstein ◽  
Yoshiko Iwamoto ◽  
Miles A. Miller ◽  
Mark A. Prytyskach ◽  
Stephane Ferretti ◽  
...  
Keyword(s):  
Cultured Cells ◽  
Temporal Dynamics ◽  
P53 Protein ◽  
Radiation Sensitivity ◽  
Negative Regulator ◽  
Normal Tissues ◽  
P53 Signaling ◽  
P53 Activation ◽  
Transcription Factor P53

AbstractRadiation sensitivity varies greatly between tissues. The transcription factor p53 mediates the response to radiation; however, the abundance of p53 protein does not correlate well with the extent of radiosensitivity across tissues. Given recent studies showing that the temporal dynamics of p53 influence the fate of cultured cells in response to irradiation, we set out to determine the dynamic behavior of p53 and its impact on radiation sensitivity in vivo. We find that radiosensitive tissues show prolonged p53 signaling after radiation, while more resistant tissues show transient p53 activation. Sustaining p53 using a small molecule (NMI801) that inhibits Mdm2, a negative regulator of p53, reduced viability in cell culture and suppressed tumor growth. Our work proposes a mechanism for the control of radiation sensitivity and suggests tools to alter the dynamics of p53 to enhance tumor clearance. Similar approaches can be used to enhance killing of cancer cells or reduce toxicity in normal tissues following genotoxic therapies.

scholarly journals TRAIL-receptor 2—a novel negative regulator of p53

2021 ◽  
Vol 12 (8) ◽  
Author(s):  
Anna Willms ◽  
Hella Schupp ◽  
Michelle Poelker ◽  
Alshaimaa Adawy ◽  
Jan Frederik Debus ◽  
...  
Keyword(s):  
Target Genes ◽  
P53 Protein ◽  
Nuclear Bodies ◽  
Negative Regulator ◽  
Trail Receptor ◽  
Wild Type ◽  
Protein Levels ◽  
Pml Nuclear Bodies ◽  
P53 Target Gene

AbstractTNF-related apoptosis-inducing ligand (TRAIL) receptor 2 (TRAIL-R2) can induce apoptosis in cancer cells upon crosslinking by TRAIL. However, TRAIL-R2 is highly expressed by many cancers suggesting pro-tumor functions. Indeed, TRAIL/TRAIL-R2 also activate pro-inflammatory pathways enhancing tumor cell invasion, migration, and proliferation. In addition, nuclear TRAIL-R2 (nTRAIL-R2) promotes malignancy by inhibiting miRNA let-7-maturation. Here, we show that TRAIL-R2 interacts with the tumor suppressor protein p53 in the nucleus, assigning a novel pro-tumor function to TRAIL-R2. Knockdown of TRAIL-R2 in p53 wild-type cells increases the half-life of p53 and the expression of its target genes, whereas its re-expression decreases p53 protein levels. Interestingly, TRAIL-R2 also interacts with promyelocytic leukemia protein (PML), a major regulator of p53 stability. PML-nuclear bodies are also the main sites of TRAIL-R2/p53 co-localization. Notably, knockdown or destruction of PML abolishes the TRAIL-R2-mediated regulation of p53 levels. In summary, our finding that nTRAIL-R2 facilitates p53 degradation and thereby negatively regulates p53 target gene expression provides insight into an oncogenic role of TRAIL-R2 in tumorigenesis that particularly manifests in p53 wild-type tumors.

scholarly journals WIP1 Inhibition by GSK2830371 Potentiates HDM201 through Enhanced p53 Phosphorylation and Activation in Liver Adenocarcinoma Cells

Cancers ◽  
2021 ◽  
Vol 13 (15) ◽  
pp. 3876
Author(s):  
Chiao-En Wu ◽  
Chen-Yang Huang ◽  
Chiao-Ping Chen ◽  
Yi-Ru Pan ◽  
John Wen-Cheng Chang ◽  
...  
Keyword(s):  
Rna Sequencing ◽  
Primary Liver Cancer ◽  
P53 Protein ◽  
Intrahepatic Bile Duct ◽  
Cell Cycle Distribution ◽  
P53 Signaling ◽  
Adenocarcinoma Cells ◽  
Wip1 Phosphatase ◽  
P53 Activation ◽  
Novel Strategy

Background: Intrahepatic cholangiocarcinoma (iCCA) is an adenocarcinoma arising from the intrahepatic bile duct. It is the second most common primary liver cancer and has a poor prognosis. Activation of p53 by targeting its negative regulators, MDM2 and WIP1, is a potential therapy for wild-type p53 cancers, but few reports for iCCA or liver adenocarcinoma exist. Methods: Both RBE and SK-Hep-1 liver adenocarcinoma cell lines were treated with the HDM201 (Siremadlin) MDM2-p53 binding antagonist alone or in combination with the GSK2830371 WIP1 phosphatase inhibitor. Cell proliferation, clonogenicity, protein and mRNA expression, cell cycle distribution, and RNA sequencing were performed to investigate the effect and mechanism of this combination. Results: GSK2830371 alone demonstrated minimal activity on proliferation and colony formation, but potentiated growth inhibition (two-fold decrease in GI50) and cytotoxicity (four-fold decrease in IC50) by HDM201 on RBE and SK-Hep-1 cells. HDM201 increased p53 protein expression, leading to transactivation of downstream targets (p21 and MDM2). Combination with GSK2830371 increased p53 phosphorylation, resulting in an increase in both p53 accumulation and p53-dependent trans-activation. G2/M arrest was observed by flow cytometry after this treatment combination. RNA sequencing identified 21 significantly up-regulated genes and five downregulated genes following p53 reactivation by HDM201 in combination with GSK2830371 at 6 h and 24 h time points compared with untreated controls. These genes were predominantly known transcriptional targets regulated by the p53 signaling pathway, indicating enhanced p53 activation as the predominant effect of this combination. Conclusion: The current study demonstrated that GSK2830371 enhanced the p53-dependent antiproliferative and cytotoxic effect of HDM201 on RBE and SK-Hep-1 cells, providing a novel strategy for potentiating the efficacy of targeting the p53 pathway in iCCA.

Sign in / Sign up

Export Citation Format

Share Document